Prevention by bombesin of cold-restraint stress induced hemorrhagic lesions in rats

Several neuropeptides, injected intraventricularly (ivt), were assessed for their effects on cold-restraint-induced hypothermia and hemorrhagic gastric lesions in 24 hr fasted rats. Bombesin (5-1 μg) further enhanced the drop in body temperature following stress and markedly prevented the gastric erosions in a dose-dependent fashion (5-0.1 μg). β-endorphin exerted a similar effect, but only at the 5 μg dose level. Other peptides (neurotensin, substance P, somatostatin and TRH: 5 μg) did not influence susceptibility to the gastric mucosal damage. Somatostatin and TRH reduced the hypothermic effect of stress. Bombesin is 250 times less potent when injected systemically than ivt and its actions are not reversed by nalaxone. The prevention of gastric erosions by bombesin could initially involve a central mechanism of action, independent of opiate receptors and possibly related to the sustained and marked hyperglycemia observed in bombesin treated rats exposed to stress.     

Unique behavioral effects of beta endorphin and their relationship to thermoregulation and hypothalamic function.

Intraventricular (ivt) injections of sub-cataleptic doses of β endorphin in rats were observed to result in wet-dog shakes. Subsequent to the wet-dog shakes, copious salivation accompanied by a clonic, seizure-like state was occasionally observed to occur. This sialogogic effect of β endorphin was blocked by naloxone and diminished by injections of thyrotropin releasing hormone. None of these behaviors were observed following ivt injection of morphine in equi-antinociceptive doses. Furthermore, hypophysectomy was observed to attenuate or block these behaviors. Both the wet-dog shakes and “sialogogic seizures” were demonstrated to be dependent upon dose as well as ambient temperature. It appears possible that the occurrence of initial wet-dog shakes may result in the elevated body temperature which then precipitates excess salivation. Since both β endorphin induced wet-dog shakes and salivation were shown to be correlated with alterations in temperature, it is possible that these effects of β endorphin indicate a physiological role for that peptide in thermoregulation.     

Effects of thyrotropin-releasing hormone on behavioral and hormonal changes induced by beta-endorphin.

Adult male rats were injected intraventricularly either with saline or TRH (10 μg) 5 min prior to a second injection of either saline or β-endorphin (50 μg). The tripeptide produced a 100% increase of motility counts recorded over a 15 min period following the last injection, whereas β-endorphin decreased general motor activity. TRH pretreatment completely abolished the depressant effect of β-endorphin. In addition, TRH enhanced the PRL secretion induced by β-endorphin and antagonized the slight elevation of plasma GH levels observed in β-endorphin-treated rats. These results do not seem to be related to an interaction of TRH with opiate receptors since the tripeptide (10^?8, 10^?6 M) added $\text{in vitro}$ to rat brain homogenates did not alter the specific binding of ³H-naloxone nor affect the displacement by β-endorphin of such binding.     

Opiate-like effects of electroconvulsive shock in rats: A differential effect of naloxone on nociceptive measures

Electroconvulsive shock (ECS) in rats produced a generalized seizure which was followed by an opiate-like catalepsy and an increase in hot-plate escape latencies. Preinjection of naloxone, at doses of 3.0 and 10.0 mg/Kg, significantly diminished the ECS-induced increase in hot-plate latencies. Paradoxically, simultaneous measurement of tail-flick latencies in these same rats demonstrated opiate-$\text{agonist}$ effects of naloxone. The cataleptic effects of ECS were demonstrated to be opiate-like by evaluating righting reflexes, grid responses, and haloperidol effects. Colonic temperatures were also measured in all animals. These data, collectively discussed relative to affective and reflexive components of nociceptive behaviors, support the hypothesis that selective endorphin systems are activated by ECS. Moreover, these observations suggest consideration of a role for endorphin systems in the therapeutic mechanisms of electroconvulsive therapy (ECT) in man.     

Humoral endorphin: A new endogenous factor with opiate-like activity

Humoral (H) endorphin, a novel endogenous opioid ligand detected in brain, blood and cerebrospinal fluid was tested in a series of opiate sensitive assays. H-endorphin displaced radiolabeled enkephalin from its specific bindings sites and inhibited the electrically evoked contraction of the guinea pig ileum and mouse vas deferens. When injected to unanesthesized animals, humoral endorphin induced analgesia in rats and mydriasis in mice. The activity of H-endorphin both $\text{in}\text{vitro}$ and $\text{in}\text{vivo}$ attests to its opioid nature. However, while its antinociceptive effect was blocked by naloxone, mydriasis induced by H-endorphin was resistant to the effect of the opiate antagonist. Similarly, intermediate concentrations of naloxone inhibited the effect of H-endorphin on the guinea pig ileum while its effect on the mouse vas deferens was completely refractory to naloxone. The physiological function of humoral endorphin in various naturally occuring states that show similar paradoxical interactions with naloxone is discussed.     

Human leukocyte interferon (HuIFN-alpha): potent endorphin-like opioid activity

Human leukocyte interferon, but not fibroblast or immune interferons, binds to opiate receptor $\text{in}\text{,}\text{vitro}$. When injected intracerebrally into mice, human leukocyte, but not fibroblast or immune interferon, caused potent endorphin-like opioid effects. These effects include analgesia, lack of spontaneous locomotion and catalepsy. All of these actions of human leukocyte interferon were preventable and reversible by the opiate antagonist naloxone. The findings suggest that some of the side effects of leukocyte interferon therapy may be mediated by opiate receptor binding. They also provide evidence for a regulatory circuit between the immune and neuroendocrine system. This putative circuit could be an etiologic site for certain psychopathological states.     

Location and characterization of opiate receptors regulating pituitary secretion

The site at which opiate agonists and antagonists act to alter secretion of prolactin, growth hormone and luteinizing hormone as well as the pharmacological specificity of the opiate receptors mediating these effects were examined in rats. Injection of β-endorphin but not a 10 fold higher dose of the non opiate peptide β-endorphin, increased release of prolactin and growth hormone in male rats while inhibiting luteinizing hormone release in ovariectomized, estrogen primed female rats. Prior treatment with naltrexone i.p. blocked these responses. Injection of naltrexone into the hypothalamus lowered prolactin release. In rats with a surgically formed hypothalamic island systemic administration of morphine or naltrexone altered prolactin release in the same manner as was observed in intact animals. In contrast no effects of β-endorphin or naltrexone were observed on the spontaneous secretion of prolactin $\text{in}$$\text{vitro}$. In addition β-endorphin did not alter the inhibition of prolactin release produced by apomorphine $\text{in}$$\text{vitro}$. The ED₅₀ for stimulation of prolactin release following intraventricular administration of β-endorphin or the synthetic enkephalin analog FK 33-824 was the same, approximately 0.1 ng/rat. However FK 33-824 at 0.2 ng/rat was able to produce much greater analgesia and catatonia than β-endorphin. The metabolism and distribution of β-endorphin was examined but did not account for these differential effects. These results indicate that opiate agonists and antagonists can act at the hypothalamic but not the anterior pituitary level to alter release of prolactin, growth hormone and luteinizing hormone. In addition the data suggest that the opiate receptors mediating release of prolactin may have a different pharmacological specificity from those involved with analgesia and catatonia.     

Interactions of thyrotropin releasing hormone and morphine sulfate in rodents.

Thyrotopin releasing hormone (TRH) produces “wet dog shakes” in rats similar to those observed during morphine withdrawal. The shaking behavior precipitated by morphine abstinence can be exacerbated by TRH administration while the other components of the morphine withdrawal syndrome remain unchanged. Morphine, chlorpromazine, apomorphine, and Δ⁹-tetrahydrocannabinol effectively block shakes induced by either TRH administration or morphine withdrawal. These results suggest the possibility that endogenous TRH may be associated with the “wet dog shakes” observed as a portion of morphine's abstinence syndrome in rats. However, TRH is unable to alter the stereospecific binding of morphine $\text{in}$$\text{vivo}$ or $\text{in}$$\text{vitro}$, and naloxone fails to potentiate the number of TRH-induced shakes. TRH has no antinociceptive properties, and it cannot alter those of morphine. These data suggest that more than one neuromechanism may be responsible for shaking behavior in rats.     

Catecholamine-induced changes in transmembrane potential and temperature in normal and dystrophic hamster brown adipose tissue

Previous studies have shown that norepinephrine (NE) elicits trans-membrane potential changes in skeletal muscle cells from normal and dystrophic (BIO 14.6) hamsters, with the magnitude of these changes being significantly less in dystrophic cells. To determine if the decreased response of the dystrophic muscle cells reflects a more generalized phenomenon, the present study was designed to evaluate the effects of NE on membrane properties of brown adipocytes. $\text{In vivo}$ techniques using glass microelectrodes were similar to those used in the muscle studies. NE injection (2 to 5 μg/kg body wt, i.v.) into anesthetized hamsters was followed by membrane depolarization, the magnitude of which did not significantly differ in the dystrophic and normal adipocytes. For example, upon administration of 5 μg NE/kg body wt, the average depolarization was $\text{14.5 ± 1.3}\text{mV}\text{(}\text{X}\text{±}\text{S.E.}\text{)}$ for 20 dystrophic cells and 14.1 ± 1.8 mV for 18 normal cells. The depolarizations following i.v. infusion of isoproterenol and phenylephrine also had similar amplitudes in both normal and dystrophic cells. Despite this lack of difference in plasma membrane responses, NE induced a significantly smaller rise in interscapular brown fat temperature in the dystrophic (0.09°C) than in the normal hamsters (0.26°C) following administration of 5 μg NE/kg body wt. Thus, the decreased responsiveness to NE of dystrophic sarcolemma did not occur with the plasma membrane of brown adipocytes, although brown fat temperature changes in the dystrophic hamsters were decreased in amplitude.     

Indentification of two somatomedin A active polypeptides and in vivo effects of a somatomedin A concentrate

The first chemical characterization of two polypeptides from human serum which stimulate the $\text{in vitro}$ incorporation of ³⁵S sulfate into chick cartilage is described. These two polypeptides, designated Somatomedin A₁ and A₂ have a molecular weight of approximately 7000. Although each peptide contains 1 cysteine residue and has asparagine as amino terminal residue, there are apparent differences in the amino acid composition. Administration of a Somatomedin A concentrate to hypophysectomized rats gave an increase in tibial width similar to that obtained with 20 μg human growth hormone.     

Morphine is most effective on gastrointestinal propulsion in rats by intraperitoneal route: Evidence for local action

A very low dose of morphine (37.5 μg/kg) entirely delivered into the peritoneal cavity of rats, consistently reduced the transit of a forced charcoal meal through the small intestine (to about 30% of control), but failed to elicit such action in naloxone-pretreated animals or if administered either intravenously or intracerebroventricularly. The same dose of tritium labeled morphine, injected i.v., resulted in brain and small intestine morphine levels respectively 2.7 times higher and 3.9 times lower than in the corresponding tissues of rats injected i.p.. These findings suggest that activation of opiate specific sites located in the gastrointestinal tract can $\text{per se}$ be primarily responsible for the antipropulsive effects of morphine.     

The 20,000-dalton structural variant of human growth hormone: lack of some early insulin-like effects

In contrast to the major form of human growth hormone the 20,000-dalton (20K) variant of the hormone produced no decrease in either serum glucose of free fatty acids one hour after injection into fasted, hypophysectomized rats. Furthermore, the variant caused no rise in serum free fatty acids after 5 hours. $\text{In}\text{vitro}$ experiments utilizing epididymal adipose tissue from hypophysectomized rats indicated that 20K was unable to accelerate glucose utilization as measured by glucose uptake and CO₂ formation. The data show that this form, even though growth promoting, lacks some of the metabolic properties attributed to growth hormone. We conclude that an insulin-like effect is not necessarily a prerequisite for growth promoting activity.     

Kyotorphin (tyrosine-arginine): Further evidence for indirect opiate receptor activation

Analgesia, opiate receptor binding, and neurochemical effects of kyotorphin (tyrosine-arginine) were studied in the rat. It was found that while kyotorphin, $\text{in vivo}$, causes naloxone reversible analgesia, and affects dopamine metabolism and acetylcholine turnover in the same manner as do morphine and other opiate agents, the dipeptide does not bind to mu, delta or kappa opiate receptors $\text{in vitro}$. Taken together, these data support the concept that there is an indirect action of kyotorphin on opiate receptors.     

Potentiation of bradykinin-induced vascular permeability increase by prostaglandin E2 and arachidonic acid in rabbit skin

The activity of prostaglandins (PG) in producing vascular permeability was quantitated by dye extraction method in skin of anaesthetized rabbits. PGE₁ and PGE₂ (0.01–10 μg) produced increase in vascular permeability. Activity was approximately equal to that of histamine (Hist) and $\text{1}\text{20}$ of that of bradykinin (BK) on a weight basis. The activity of PGF_1α and PGF_2α was only $\text{1}\text{20}$ of that of PGE₁ or PGE₂.In spite of the relatively low potency of PGE₁ and PGE₂ in the rabbit, near threshold doses (0.1 or 1 μg) of PGE₂ could potentiate permeability responses to bradykinin (0.1 μg) by 10 or 100-fold, respectively. Equivalent doses (0.1 or 1 μg) of histamine could not potentiate the bradykinin responses. Arachidonic acid (AA) at 1 μg, produced a 10-fold potentiation in the permeability response to bradykinin (0.1 μg). Pretreatment of the rabbits with indomethacin (20 mg/kg, i.p.) reduced the responses of BK (0.1 μg) + AA (1 μg) down to a similar magnitude of those seen with bradykinin alone. However, indomethacin did not block responses to either, BK alone, BK + PGE₂, or BK + Hist. Various doses (1, 10, 100 and 300 μg) of arachidonic acid alone also produced increase in cutaneous vascular permeability, although its potency was only $\text{1}\text{3}$–$\text{1}\text{8}$ of that of PGE₂. This activity of arachidonic acid was attributed in part to its bioconversion to PGE₂, since its activity was significantly reduced by the prostaglandin antagonist, diphloretin phosphate (DPP) (60 mg/kg, i.v.) and by indomethacin (20 mg/kg, i.p.), which blocks conversion of arachidonic acid to prostaglandins. Arachidonic acid may owe some of its permeability increaseing effects to histamine release, since its effects were also reduced by the antihistamine, pyrilamine (2.5 mg/kg, i.v.).     

Stimulation of leydig cell testoterone secretion in vitro and in vivo in hypophysectomized rats by an agonist of luteinizing hormone releasing hormone

Injection of a luteinizing hormone-releasing hormone (LHRH) agonist into 55-day-old male rats which had been hypophysectomized 3 days earlier resulted in a 10- to 30-fold increase in the levels of testosterone in serum and testicular interstitial fluid (IF) in the 4h following injection. The levels achieved were within or above the normal range for intact untreated rats of this age. In similar animals, injection of LHRH agonist also enhanced the serum testosterone response to injected hCG at $\text{1}\text{1}\text{2}\text{h}$, but not at later times after injection, and by 24h reduced IF levels of testosterone suggested that LHRH agonist had begun to inhibit stimulation by hCG. $\text{In vitro}$, dispersed Leydig cells from untreated hypophysectomized rats showed a 2-fold increase in testosterone responsiveness to LHRH agonist when compared to cells from intact rats, and this change was associated with an 80% increase in the number of Leydig cell LHRH-receptors.     

Evidence that opiate receptors mediate suppression of hypertonic saline-induced drinking in the mouse by narcotic antagonists

The effects of naloxone, its $\text{dextro}$-stereisomer, and five other narcotic antagonists were determined on water intake induced by intracellular dehydration in the mouse. The intraperitoneal administration of a 2M sodium chloride solution served as the model for intracellular dehydration. $\text{1}$-Naloxone (0.01-10 mg/kg) reduced drinking in a dose-dependent fashion with an ED₅₀ of 0.55 mg/kg. In contrast, $\text{d}$-naloxone failed to suppress water consumption at doses up to 10 mg/kg. The other narcotic antagonists tested --- naltrexone, diprenorphine, levallorphan, oxilorphan, and nalorphine --- also produced dose-dependent decreases in water consumption. The order of potency of these narcotic antagonists in suppressing water intake was highly correlated with their orders of potency in other procedures involving the opiate receptor. The stereoselectivity and order of potency suggest that the suppressant effects of the narcotic antagonists on drinking induced by hypertonic saline administration in the mouse are mediated through an opiate receptor-dependent mechanism.     

Substance P found to lower body temperature and aggression.

Synthetic substance P (SP) was bioassayed in mice by a procedure comprising eleven subtests. Two replications of a dose-response study were conducted at the time of the peak effect of the intravenous injection of SP. Single doses of 31 and 63 ng/kg significantly decreased body temperature. SP, [D-Arg¹]-SP, and [des-NH₂-Arg¹]-SP comparably lowered blood pressure, but [D-Arg¹]-SP and [des-NH₂-Arg¹]-SP were $\text{1}\text{20}$ to $\text{1}\text{10}$ as active as SP in lowering body temperature. The activities that lower body temperature and blood pressure may be different. The thyrotropin and luteinizing hormone releasing hormones (TRH and LH-RH) did not lower body temperature. SP also decreased the aggressive response (ED₅₀, 89 ng/kg).     

Onset of the receptive and proceptive components of feminine sexual behavior in rats following the intravenous administration of progesterone

The present study was carried out in order to assess the time course of action of progesterone (P) in the facilitation of complete feminine sexual behavior. Female rats (estrogen primed via 5% E₂ Silastic capsules) were given 200 μg of P either intravenously (iv) or subcutaneously (sc), and tested for estrous behavior at $\text{1}\text{4}$, $\text{1}\text{2}$, 1, 2, and 4 hr after treatment. Among iv-treated animals, significant amounts of lordosis behavior were seen as early as $\text{1}\text{2}$ hr, and a dramatic rise in solicitation behavior was observed at 2 hr. Although sc-treated animals displayed significant amounts of lordosis and solicitation behavior at 2 hr, the behavior was not maximal until 4 hr. Intravenous administration of 400 μg P was equipotent to 200 μg P, whereas 50 μg of iv P was relatively ineffective. A dual mechanism hypothesis pertaining to progesterone's actions in the facilitation of both the receptive and preceptive components of feminine sexual behavior in rats is discussed.     

Bacteria-derived human growth hormone lacks lipolytic activity in rat adipose tissue

Bacteria-derived human growth hormone (hGH) shows little $\text{in}\text{vitro}$ lipolytic activity in adipose tissue from fed rats. In adipose tissue from fasted rats no lipolytic activity is observed. However, bacteria-derived hGH increased serum free fatty acids after intraperitoneal administration to hypophysectomized rats to the same extent as purified pituitary hGH. The dose response of the bacteria-derived hGH tested for $\text{in}\text{vitro}$ insulin-like activity was very similar to the pituitary extracted material. Thus bacteria-derived hGH behaves in a manner indistinguishable from highly purified preparations of pituitary hGH.     

Monitoring the stereospecificity of morphine action in vivo and in vitro through brain membrane lipid fluidity

Differential scanning calorimetry of crude brain mitochondrial lipids obtained from control and morphine treated rats was carried out and the lipid phase transition measured. Morphine treatment resulted in a significant decrease in the temperature range and enthalpy of the phase transition. This effect was found to be dose dependent and reversible both $\text{in vivo}$ and $\text{in vitro}$ by naloxone. Studies with levorphanol and dextrorphan demonstrated stereospecificity. Furthermore, the ether precipitable fraction of total lipid extracts was shown to mediate the opiate response.